There are a group of proteins that constitute the eukaryotic protein kinase superfamily. Enzymes of this class specifically phosphorylate serine, threonine or tyrosine residues of intracellular proteins. These enzymes are important in mediating signal transduction in multicellular organisms. Many of the protein kinases are part of transmembrane receptors. Others occur as intracellular proteins which take part in signal transduction within the cell, including signal transduction to the nucleus and activation of other proteins. Other protein kinases, such as G protein-coupled receptor kinases, are bound to cell membranes and participate in transmembrane signaling.
As such, phosphorylation of serine, threonine or tyrosine by protein kinases is an important mechanism for regulating intracellular events in response to environmental changes. A wide variety of cellular events are regulated by protein kinases. A few examples include cellular proliferation, cellular differentiation, the ability of cells to enter and/or complete mitosis, cellular transformation by RNA viruses, oncogenesis, control of fat metabolism, immune responses, inflammatory responses and the control of carbohydrate metabolism.
Enhanced protein kinase activity can lead to persistent stimulation by secreted growth factors and other growth inducing factors which, in turn, can lead to proliferative diseases such as cancer, to nonmalignant proliferative diseases such as arteriosclerosis, psoriasis and to inflammatory response such as septic shock. Decreased function can also lead to disease. For example, a decrease in the activity of insulin receptor kinase is a cause of various types of diabetes. Severe reduction of the B cell progenitor kinase leads to human X-linked agammaglobulinemia.
Thus, agents which can modulate (increase or decrease) the activity of protein kinases have great potential for the treatment of a wide variety of diseases and conditions such as cancer, obesity, autoimmune disorders, inflammation and diabetes. Such agents also have utility in deciphering the mode of action of protein kinases and how these proteins regulate cellular functions and activities.
It has now been found that short peptides which are derivatives of the xcex1D region of a protein kinase can significantly affect the activities of cells expressing the protein kinase when incubated with the cells (the xe2x80x9cxcex1D regionxe2x80x9d is defined hereinbelow). For example, the peptide derivatives of the xcex1D region of Jak3 inhibit the proliferation of human endothelial cells and the human prostate cancer cell line PC3 in vitro at concentrations as low as 0.3 xcexcM (Example 2). Based on the aforementioned discoveries, novel peptides are disclosed herein which are peptide derivatives of the xcex1D region of protein kinases. Also disclosed are methods of identifying a peptide derivative of an xcex1D region of a protein kinase that modulates the activity of the protein kinase. Methods of modulating the activity of a protein kinase in a subject are also disclosed.
One embodiment of the present invention is a novel peptide which is a peptide derivative of the xcex1D region of a protein kinase. The peptide comprises between about five and about thirty amino acid residues or amino acid residue analogs of the xcex1D region. The peptide modulates the activity of the protein kinase. The N-terminus and/or C-terminus of the peptide can be substituted or unsubstituted. The peptide can be linear or cyclic.
Another embodiment of the present invention is a method of modulating the activity of a protein kinase in a subject. The method comprises administering a therapeutically effective amount of a peptide that is a derivative of the xcex1D region of the protein kinase, as described above.
Yet another embodiment of the present invention is a method of identifying a peptide which modulates the activity of a protein kinase. The method comprises providing a xe2x80x9ctest peptidexe2x80x9d which has from about five to about thirty amino acids or amino acid analogs and which is a peptide derivative of the xcex1D region of the protein kinase. The test peptide is incubated with cells having a cellular activity or function under the control of the protein kinase under conditions suitable for assessing the activity of the protein kinase. The activity of the protein kinase is assessed and compared with the activity of the protein kinase in cells of the same cell type grown under the same conditions in the absence of the test peptide. A greater or lesser activity compared with cells grown in the absence of the test peptide indicates that the test peptide modulates the activity of the protein kinase.
The peptides of the present invention can be used in the treatment of a wide variety of diseases caused by overactivity or underactivity of a protein kinase. Examples include, but are not limited to, cancer, diseases caused by proliferation of smooth muscle (e.g., restenosis and atherosclerosis), skin disorders, diabetes, obesity, diseases of the central nervous system, inflammatory disorders, autoimmune diseases and other immune disorders, osteoporosis and cardiovascular diseases. The peptides of the present invention also have in vitro utilities, for example, in the generation of antibodies that specifically bind the protein kinase from which the peptide was derived. These antibodies can be used to identify cells expressing the protein kinase and to study the intracellular distribution of the protein kinase. In addition, the peptides of the present invention can be used to identity and quantitate ligands that bind the xcex1D region of the protein kinase from which the peptide was derived.